Peripheral nerve injury is known to induce a variety of molecular and physiological changes in primary sensory and dorsal horn neurons. Target derived neurotrophic signaling may play an important role in the injury response. The long-term goal of these studies is to determine how changes in the afferent target regions alter the response properties of dorsal root ganglion (DRG) neurons after nerve injury. We first propose to test the hypothesis that nerve crush and regeneration of saphenous afferents induces altered levels and localization of various neurotrophic factors and their receptors in the skin and DRGs, respectively, over time. Our preliminary evidence suggests that peripheral axotomy induces changes in several neurotrophic factor levels, including NGF and artemin, in the skin, and this correlates to increases in receptors like TRPV1 in the DRGs. In turn, this may underlie our recent finding that nerve injury results in changes in the percentage of TRPV1 containing C-fiber neurons that only respond to heat (CH) after regeneration. For example, high levels of NGF regulate the expression of TRPV1 in dissociated/ axotomized DRG neurons in vitro, and overexpression of artemin in the skin enhances TRPV1 mRNA in the DRGs in vivo. Given that TRPV1, NGF receptor trkA and artemin receptor GFRaS immunostaining overlap extensively in the DRG and each have been linked to pain responses, we then propose to test if GFRaS and/or trkA mediated neurotrophic factor signaling regulates the injury induced changes in the CH neurons. We will be able to correlate the known response properties of regenerated sensory neurons to changes in neurotrophic factor signaling immunocytochemically, and using RT-PCR and western blots before and after reinnervation of the skin, and we will be able to functionally test the role of two particular pathways in establishing the changes in electrophysiological response properties of CH neurons after injury using an ex vivo skin-nerve-DRG-spinal cord preparation after in vivo siRNA mediated knockdown of GFRaS and/or trkA. These studies will allow us to further understand the functional implications of excess growth factor signaling in the skin on injured neurons and how these changes relate to nerve injury induced hyperalgesia. This work may also lead us to better ways to treat injury related chronic pain states.